Abstract

Background

The zebrafish embryo is an important in vivo model to study the host innate immune response towards microbial infection. In most
zebrafish infectious disease models, infection is achieved by micro-injection of bacteria
into the embryo. Alternatively, Edwardsiella tarda, a natural fish pathogen, has been used to treat embryos by static immersion. In
this study we used transcriptome profiling and quantitative RT-PCR to analyze the
immune response induced by E. tarda immersion and injection.

Results

Mortality rates after static immersion of embryos in E. tarda suspension varied between 25-75%, while intravenous injection of bacteria resulted
in 100% mortality. Quantitative RT-PCR analysis on the level of single embryos showed
that expression of the proinflammatory marker genes il1b and mmp9 was induced only in some embryos that were exposed to E. tarda in the immersion system, whereas intravenous injection of E. tarda led to il1b and mmp9 induction in all embryos. In addition, microarray expression profiles of embryos subjected
to immersion or injection showed little overlap. E. tarda-injected embryos displayed strong induction of inflammatory and defense genes and
of regulatory genes of the immune response. E. tarda-immersed embryos showed transient induction of the cytochrome P450 gene cyp1a. This gene was also induced after immersion in Escherichia coli and Pseudomonas aeruginosa suspensions, but, in contrast, was not induced upon intravenous E. tarda injection. One of the rare common responses in the immersion and injection systems
was induction of irg1l, a homolog of a murine immunoresponsive gene of unknown function.

Conclusions

Based on the differences in mortality rates between experiments and gene expression
profiles of individual embryos we conclude that zebrafish embryos cannot be reproducibly
infected by exposure to E. tarda in the immersion system. Induction of il1b and mmp9 was consistently observed in embryos that had been systemically infected by intravenous
injection, while the early transcriptional induction of cyp1a and irg1l in the immersion system may reflect an epithelial or other tissue response towards
cell membrane or other molecules that are shed or released by bacteria. Our microarray
expression data provide a useful reference for future analysis of signal transduction
pathways underlying the systemic innate immune response versus those underlying responses
to external bacteria and secreted virulence factors and toxins.